Different blocking effects of HgCl2 and NaCl on aquaporins of pepper plants

In this study we have compared the short-term effects of both NaCl and HgCl₂ on aquaporins of Capsicum annuum L. plants, in order to determine whether or not they are similar. Stomatal conductance, turgor, root hydraulic conductance and water status were measured after 0.5, 2, 4 and 6 h of NaCl (60 mmol/L) or HgCl₂ (50 μmol/L) treatment. When 60 mmol/L NaCl was added to the nutrient solution, a large decrease in stomatal conductance was observed after 2 h. However, when HgCl₂ (50 μmol/L) was added, the decrease occurred after 4 h. The number of open stomata closed was always lower in plants treated with HgCl₂ than in plants treated with NaCl. The water content of the Hg²⁺-treated plants was decreased, compared with controls and NaCl-treated. The root hydraulic conductance decreased after HgCl₂ and NaCl treatment plants. Turgor of leaf epidermal cells was greatly reduced in plants treated with HgCl₂, but remained constant in the NaCl treatment, compared with control plants. The fact that the stomatal conductance was reduced more rapidly after NaCl addition, followed by the stomatal closure, and that both water content and turgor did not differ from the control suggests that in NaCl-treated plants there must be a signal moving from root to shoot. Therefore, the control of plant homeostasis through a combined regulation of root and stomatal exchanges may be dependent on aquaporin regulation.     

Baclofen reestablishes striatal and cortical dopamine concentrations during naloxone-precipitated withdrawal

The present study analyzes the effects of baclofen (BAC) on mice brain neurochemical alterations during the morphine (MOR) withdrawal syndrome. Male Swiss-Webster albino mice (27-33 g) were rendered dependent by intraperitoneal (i.p.) injection of MOR (2mg/kg), twice daily for 9 days. On day 10, the dependent animals were divided into two groups: one receiving naloxone (NAL; 6 mg/kg i.p.) to precipitate the withdrawal syndrome 60 min after the last dose of MOR and the other received BAC (2mg/kg, i.p.) followed by NAL (6 mg/kg, i.p.), injected 30 and 60 min after the last dose of MOR, respectively. Ten minutes after these treatments, mice were killed by decapitation and the striatum, cortex and hippocampus were dissected to determine endogenous concentrations of dopamine (DA), 5-hydroxytryptamine (5-HT) and their metabolites using HPLC with electrochemical detection. Striatal DA, dihydroxyphenyl acetic acid (DOPAC) and homovanillic acid (HVA) concentrations as well as cortical DA concentrations of the withdrawal groups decreased significantly with respect to the control groups. BAC attenuated the decrease in DA and DOPAC concentrations observed during the withdrawal, without modifying per se the control DA concentrations. No changes on 5-HT and its metabolite 5-hydroxyindolacetic acid (5-HIAA) concentrations were observed during the MOR abstinence syndrome. The prevention caused by BAC on the decreased concentrations of DA induced by MOR withdrawal could have a therapeutic interest for the management of withdrawal syndrome.     

Prostaglandin f(2alpha) induces distinct physiological responses in porcine corpora lutea after acquisition of luteolytic capacity

This study examines differences in intracellular responses to cloprostenol, a prostaglandin (PG)F(2alpha) analog, in porcine corpora lutea (CL) before (Day 9 of estrous cycle) and after (Day 17 of pseudopregnancy) acquisition of luteolytic capacity. Pigs on Day 9 or Day 17 were treated with saline or 500 microgram cloprostenol, and CL were collected 10 h (experiment I) or 0.5 h (experiment III) after treatment. Some CL were cut into small pieces and cultured to measure progesterone and PGF(2alpha) secretion. In experiment I, progesterone remained high and PGF(2alpha) low in luteal incubations from either Day 9 or Day 17 saline-treated pigs. Cloprostenol increased PGF(2alpha) production 465% and decreased progesterone production 87% only from Day 17 luteal tissue. Cloprostenol induced prostaglandin G/H synthase (PGHS)-2 mRNA (0.5 h) and protein (10 h) in both groups. In cell culture, cloprostenol or phorbol 12, 13-didecanoate (PDD) (protein kinase C activator), induced PGHS-2 mRNA in luteal cells from both groups. However, acute cloprostenol treatment (10 min) decreased progesterone production and increased PGF(2alpha) production only from Day 17 luteal cells. Thus, PGF(2alpha) production is induced by cloprostenol in porcine CL with luteolytic capacity (Day 17) but not in CL without luteolytic capacity (Day 9). However, this change in PGF(2alpha) production is not explained by a difference in induction of PGHS-2 mRNA or protein.     

Sequence and diversity of MHC <Emphasis Type="Italic">DQA</Emphasis> and<Emphasis Type="Italic"> DQB</Emphasis> genes of the owl monkey<Emphasis Type="Italic"> Aotus nancymaae</Emphasis>

The New World primate Aotus nancymaae has been recommended by the World Health Organization (WHO) as a model for evaluation of malaria vaccine candidates, given its susceptibility to experimental infection with the human malaria parasites Plasmodium falciparum and Plasmodium vivax. We present here the nucleotide sequences of the complete cDNA of MHC-DQA1 and of the polymorphic exon 2 segments of MHC-DQB1/DQB2. In a group of three nonrelated animals captured in the wild, five alleles of MHC-DQA1 could be identified. They all belong to one lineage, namely Aona-DQA1*27. This lineage has not been described in any other New World monkey species studied. In a group of 19 unrelated animals, 14 Aona-DQB1 alleles could be identified which are grouped into the two lineages Aona-DQB1*22 and Aona-DQB1*23. These lineages have been described previously in the common marmoset and cotton-top tamarin. In addition, two Aona-DQB2 sequences could be identified which are highly similar to HLA-DQB2 sequences. Essential amino acid residues contributing to MHC DQ peptide binding pockets number 1 and 4 are conserved or semi-conserved between HLA-DQ and Aona-DQ molecules, indicating a capacity to bind similar peptide repertoires. These results fully support the use of Aotus monkeys as an animal model for evaluation of future subunit vaccine candidates.     

Cloning of an endangered species (Bos gaurus) using interspecies nuclear transfer

Approximately 100 species become extinct a day. Despite increasing interest in using cloning to rescue endangered species, successful interspecies nuclear transfer has not been previously described, and only a few reports of in vitro embryo formation exist. Here we show that interspecies nuclear transfer can be used to clone an endangered species with normal karyotypic and phenotypic development through implantation and the late stages of fetal growth. Somatic cells from a gaur bull (Bos gaurus), a large wild ox on the verge of extinction, (Species Survival Plan < 100 animals) were electrofused with enucleated oocytes from domestic cows. Twelve percent of the reconstructed oocytes developed to the blastocyst stage, and 18% of these embryos developed to the fetal stage when transferred to surrogate mothers. Three of the fetuses were electively removed at days 46 to 54 of gestation, and two continued gestation longer than 180 (ongoing) and 200 days, respectively. Microsatellite marker and cytogenetic analyses confirmed that the nuclear genome of the cloned animals was gaurus in origin. The gaur nuclei were shown to direct normal fetal development, with differentiation into complex tissue and organs, even though the mitochondrial DNA (mtDNA) within all the tissue types evaluated was derived exclusively from the recipient bovine oocytes. These results suggest that somatic cell cloning methods could be used to restore endangered, or even extinct, species and populations.     

ADP modulates the dynamic behavior of the glycolytic pathway of Escherichia coli

A mathematical model that includes biochemical interactions among the PTS system, phosphofructokinase (PFK), and pyruvate kinase (PK) is used to evaluate the dynamic behavior of the glycolytic pathway of Escherichia coli under steady-state conditions. The influence of ADP, phosphoenolpyruvate (PEP), and fructose-6-phosphate (F6P) on the dynamic regulation of this pathway is also analyzed. The model shows that the dynamic behavior of the system is affected significantly depending on whether ADP, PEP, or F6P is considered constant a steady state. Sustained oscillations are observed only when dADP/dt not equal 0 and completely suppressed if dADP/dt = 0 at any steady-state value. However, when PEP or F6P is constant, the system evolves toward the formation of stable limit cycles with periods ranging from 0.2 min to hours.     

Prostaglandin E(2) increases bovine leukemia virus tax and pol mRNA levels via cyclooxygenase 2: regulation by interleukin-2, interleukin-10, and bovine leukemia virus

Prostaglandin E(2) (PGE(2)), produced by macrophages, has important immune regulatory functions, suppressing a type 1 immune response and stimulating a type 2 immune response. Type 1 cytokines (interleukin-2 [IL-2], IL-12, and gamma interferon) increase in freshly isolated peripheral blood mononuclear cells (PBMCs) of animals with an early disease stage of bovine leukemia virus (BLV) infection, while IL-10 increases in animals with a late disease stage. Although IL-10 has an immunosuppressive role in the host immune system, IL-10 also inhibits BLV tax and pol mRNA levels in vitro. In contrast, IL-2 stimulates BLV tax and pol mRNA and p24 protein expression in cultured PBMCs. The inhibitory effect of IL-10 on BLV expression depends on soluble factors secreted by macrophages. Thus, we hypothesized that PGE(2), a cyclooxygenase 2 (COX-2) product of macrophages, may regulate BLV expression. Here, we show that the level of COX-2 mRNA was decreased in PBMCs treated with IL-10, while IL-2 enhanced the level of COX-2 mRNA. Addition of PGE(2) stimulated BLV tax and pol mRNA levels and reversed the IL-10 inhibition of BLV mRNA. In addition, the specific COX-2 inhibitor, NS-398, inhibited the amount of BLV mRNA detected. Addition of PGE(2) increased BLV tax mRNA regardless of NS-398 addition. PGE(2) inhibited antigen-specific PBMC stimulation, suggesting that stimulation of BLV tax and pol mRNA levels by PGE(2) is independent of cell proliferation. These findings suggest that macrophage-derived COX-2 products, such as PGE(2), regulate virus expression and disease progression in BLV infection.     

EPR characterization of mono(thiosemicarbazones) copper(II) complexes. Note II

Copper(II) complexes with thiosemicarbazones have been shown to be more active in cell destruction, in the inhibition of DNA synthesis than the uncomplexed ligand. Several derivatives of thiosemicarbazones and their iron and copper complexes have been studied for their cytotoxicity and inhibiting activity against DNA synthesis. In the present work complexes formed in H2O-DMSO solution between copper(II) and the acetophenone thiosemicarbazone (ATSC) and the o-aminobenzaldehyde thiosemicarbazone (o-NH2TSC) have been studied. EPR studies have been performed at different pH values and metal-to-ligand ratios. The spectra have been recorded at both room (298 K) and low temperatures (120 K). A possible relationship between structure and activity is attempted on the basis of the EPR data.     

Role of Potassium Channels in Amyloid-Induced Cell Death

Abstract: Basal forebrain cholinergic neurons are severely depleted early in Alzheimer's disease and appear particularly susceptible to amyloid β-peptide (Aβ) toxicity in vivo. To model this effect in vitro, a cholinergic septal cell line (SN56) was exposed to Aβ. SN56 cells exhibited a tetraethylammonium (TEA)-sensitive outward K⁺ current with delayed rectifier characteristics. Increases of 64% (±19; p < 0.02) and 44% (±12; p < 0.02) in K⁺ current density were noted 6–12 and 12–18 h following the addition of Aβ to SN56 cell cultures, respectively. Morphological observation and staining for cell viability showed that 25 ± 4 and 39 ± 4% of SN56 cells were dead after 48- and 96-h exposures to Aβ, respectively. Perfusion of SN56 cells with 10–20 mM TEA blocked 71 ± 6 to 92 ± 2% of the outward currents, widened action potentials, elevated [Ca²⁺]_i, and inhibited 89 ± 14 and 68 ± 14% of the Aβ toxicity. High [K⁺]_o, which depolarizes cell membranes and increases [Ca²⁺]_i, also protected SN56 cells from Aβ toxicity. This effect appeared specific since glucose deprivation of SN56 cells did not alter K⁺ current density and TEA did not protect these cells from hypoglycemic cell death. Furthermore, Aβ was toxic to a dopaminergic cell line (MES23.5) that expressed a K⁺ current with delayed rectifier characteristics; K⁺ current density was not altered by Aβ and MES23.5 cells were not protected by TEA from Aβ toxicity. In contrast, a noncholinergic septal cell line (SN48) that shows minimal outward K⁺ currents was resistant to the toxicity of Aβ. These data suggest that a K⁺ channel with delayed rectifier characteristics may play an important role in Aβ-mediated toxicity for septal cholinergic cells.